Este trabalho é apoiado por uma bolsa do Conselho de Pesquisa da Universidade de Baylor e de uma bolsa de pesquisa da Fundação de Epilepsia.

Os ratos utilizados no experimento seguinte foram gerados e alojados na Universidade de Baylor em uma temperatura ambiente de 22 ° C, com uma luz de 14 horas e 10 horas escuro (20:00-06:00 hr) ciclo diurno. Os ratinhos foram fornecidas ad libitum acesso a comida e água. Todos os procedimentos para os ratos estavam em conformidade com os Institutos Nacionais de Saúde Diretrizes para o Cuidado e Uso de Animais de Laboratório e do protocolo de animais foi aprovado pela Universidade de Baylor Animal Care e do Comitê Use. Visão global A tarefa do medo condicionado traço é baseado em procedimentos descritos por Wiltgen e colegas 9. 1. Preparação do Equipamento A câmara de aparelho de condicionamento do medo (26 centímetros x 22 cm x 18) consiste de duas partes que são de acrílico, de dois lados, que são de metal, e um chão de grade de fundo, que é usado para fornecer um choque no pé leve. A câmara de teste é a casad em uma câmara de som atenuado. A câmara também é apertado luz para evitar a luz do lado de fora de influenciar o software de detecção de movimento. <ol><li> Calibrar os níveis de choque, os níveis de luz, e os níveis de intensidade sonora para a câmara de testes. Medir o nível de fundo da câmara de testes. O ruído de fundo nesta câmara é de 65 dB. Use um medidor de som para medir este nível. O medidor deve ser ajustado para 70 dB, definido como C, e para a detecção de ajuste lento. <ol><li> Calibrar o nível de choque de 0,5 mA. Use um dispositivo de calibração externa para calibrar corretamente o nível de choque (ver Tabela Materiais). A medição interna do gerador de choque não é preciso. O gerador de choque administra um choque mexidos que não pode ser medido com precisão por um medidor de amp padrão. </li><li> Coloque uma das pontas em um dos bares da rede e colocar os outros levam 3 ou 4 bares mais. Use o gerador de choque para administrar o choque. Ajuste o nível de choque até o nível correto seja atingido. Faça isso por eAch temer câmara de condicionamento. </li><li> Feche a porta para a câmara quando se mede o nível de luz da câmara. Calibrar o nível de luz de aproximadamente 1,0. Este é um número específico para o software freezeframe utilizado nesta experiência. Um medidor de luz externa vai ler isso como 2 lux. O nível de luz pode ser ajustada movendo a localização da luz casa ou ajustando o condensador da lente. Certifique-se de apertar os parafusos de ajuste para a lente após o ajuste. </li><li> Calibrar o nível do som em decibéis a 85 dB. Use um decibelímetro externo dentro da gaiola de testes para calibrar o nível de decibéis (ver Tabela Materiais). O som apresentado será um tom Hz 2.700. Nota: Se estiver usando camundongos com idade superior a 6 meses de idade, pode ser melhor usar o ruído branco desde ratos mais velhos podem ter déficit auditivo. </li></ol></li><li> Depois que o aparelho está pronto levar os sujeitos a uma sala de espera separada. Nota: Não abrigar o mouse na mesma sala, como a sala de testes. </li><li> ª rótuloe a cauda dos ratos que serão testadas em ordem de teste. É melhor reduzir o manuseio excessivo antes do teste. Alternativamente, as caudas podem ser rotuladas no dia anterior ao ensaio para reduzir a lidar com o stress. Após os ratinhos foram rotulados permitir a aclimatar ao ambiente durante 30 minutos. Tenha gaiolas adicional limpas para abrigar os ratos após o teste é concluído. </li></ol> 2. Traçar Condicionado Dia 1 <ol><li> Retire cada rato da gaiola e colocá-los em gaiolas individuais para o transporte para a câmara de medo condicionado. Use roupa de cama limpa para cada gaiola. Coloque uma nota em gaiola transferência para manter a ordem de camundongos testando correta. Nota: Se os ratos são alojados individualmente em seguida, eles podem ser transportadas na sua gaiola de origem. </li><li> Posicione o mouse na câmara de teste e fechar a porta. Inicie o programa de software. </li><li> No dia de treino, permite que os ratinhos para explorar a câmara durante 3 min. O software em seguida, apresenta a 20 seg de tom (85 dB, 2.700 Hz) to animal. Depois de um período de rastreio de 20 seg um choque ligeiro (2 s, 0,5 mA) é administrado ao animal. </li><li> Registre a reação do assunto para confirmar que recebeu os estímulos aversivos, visualizando o vídeo. Um intervalo inter-julgamento 200 seg separa os ensaios 5 condicionamento. Cada ensaio consiste de 20 seg tom seguido por um atraso de 20 segundos, em seguida, um choque. </li><li> Após o teste é concluído que os animais possam permanecer na câmara de teste durante 1 minuto antes de a retirar da gaiola de teste. </li><li> Colocar o animal de volta na gaiola de transferência e devolvê-lo à sua gaiola. Se houver ratos adicionais em sua gaiola casa, então abrigar individualmente o mouse até que todos os ratos de teste completo. Isto irá reduzir a tensão para o outro ratos que não foram testados. Uma solução alternativa seria a de abrigar individualmente todos os ratinhos uma semana antes do ensaio, para reduzir a influência da remoção contínua ratinhos de uma gaiola. </li><li> Limpe a câmara de testes com 30% de isopropanol após each animal é testado. </li><li> Repita os passos de 2,2-2,7 para todos os murganhos no grupo do teste. </li><li> Retornar todos os ratos para o quarto colônia após o último rato na coorte foi testado. </li></ol> 3. Rastreamento Medo Condicionado Day 2: Teste de memória Rastreamento <ol><li> Teste de memória rastreamento ocorrerá no dia 2. Neste protocolo há 3 apresentações de tom. Coloque os ratos em um novo contexto para o teste traço condicionado. </li><li> Prepare o software para executar um programa com um período inicial de 2 min seguido por três apresentações de 20 segundos de tom. Há sa 220 sec ITI entre cada apresentação tom. </li><li> Para a nova condição contexto, colocar inserções de acrílico no chão da câmara para alterar a forma, textura e cor da câmara de condicionamento. </li><li> Alterar o odor na câmara, colocando extrato de baunilha em um barco pesar sob a inserção chão. </li><li> Limpar a câmara com etanol a 70% em vez de 30% de isopropanol. Nota: Isso vai ajudarpara criar um novo contexto. </li><li> Traga os ratos para a sala de detenção e relabel suas caudas para testes, se necessário. </li><li> Prepare novas gaiolas de transferência contexto, substituindo a cama com papel picado. Nota: Isso vai ajudar na criação de um novo contexto. </li><li> Coloque os ratos na câmara de teste, em seguida, iniciar o programa. Limpar a câmara com etanol a 70% após o teste é concluído. </li><li> Retorne os ratos para sua gaiola para casa após o traço condicionado está concluída. Retornar todos os ratos para o quarto colônia quando todos os ratos foram testados. </li></ol> 4. Rastreamento Medo Condicionado Dia 3: Teste de memória Contextual <ol><li> No terceiro dia de condicionamento contextual é conduzida. Prepare o software para executar um programa para gravar o comportamento de congelamento por 8 min. </li><li> Limpe a câmara com 30% isopropanol antes de testar e depois de testar cada rato. O contexto deve ser idêntica à do dia 1. As gaiolas de transferência deve ser o mesmo que no dia 1. </li><li&gt; Trazer camundongos para a sala de detenção e rotular suas caudas para testes. </li><li> Coloque cada rato na câmara de teste, em seguida, iniciar o programa. Limpe a câmara com 30% de isopropanol após o teste é concluído. </li><li> Retornar todos os ratos para o quarto colônia quando concluído. </li></ol>

<ol>
	<li>Powell, R. A., Honey, P. L., Symbaluk, D. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Introduction to learning and behavior. , .</li><li>Tsaltas, E., Preston, G. C., Gray, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effects+of+dorsal+bundle+lesions+on+serial+and+trace+conditioning.">The effects of dorsal bundle lesions on serial and trace conditioning.</a> Behav. Brain Res. 10, 361-374 (1983).</li><li>McAlonan, G. M., Dawson, G. R., Wilkinson, L. O., Robbins, T. W., Everitt, B. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effects+of+AMPA-induced+lesions+of+the+medial+septum+and+vertical+limb+nucleus+of+the+diagonal+band+of+Broca+on+spatial+delayed+non-matching+to+sample+and+spatial+learning+in+the+water+maze.">The effects of AMPA-induced lesions of the medial septum and vertical limb nucleus of the diagonal band of Broca on spatial delayed non-matching to sample and spatial learning in the water maze.</a> Eur. J. Neurosci. 7, 1034-1049 (1995).</li><li>Chowdhury, N., Quinn, J. J., Fanselow, M. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dorsal+hippocampus+involvement+in+trace+fear+conditioning+with+long,+but+not+short,+trace+intervals+in+mice.">Dorsal hippocampus involvement in trace fear conditioning with long, but not short, trace intervals in mice.</a> Behav. Neurosci. 119, 1396-1402 (2005).</li><li>Quinn, J. J., Oommen, S. S., Morrison, G. E., Fanselow, M. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Post-training+excitotoxic+lesions+of+the+dorsal+hippocampus+attenuate+forward+trace,+backward+trace,+and+delay+fear+conditioning+in+a+temporally+specific+manner.">Post-training excitotoxic lesions of the dorsal hippocampus attenuate forward trace, backward trace, and delay fear conditioning in a temporally specific manner.</a> Hippocampus. 12, 495-504 (2002).</li><li>Misane, I., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Time-dependent+involvement+of+the+dorsal+hippocampus+in+trace+fear+conditioning+in+mice.">Time-dependent involvement of the dorsal hippocampus in trace fear conditioning in mice.</a> Hippocampus. 15, 418-426 (2005).</li><li>Quinn, J. J., Loya, F., Ma, Q. D., Fanselow, M. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dorsal+hippocampus+NMDA+receptors+differentially+mediate+trace+and+contextual+fear+conditioning.">Dorsal hippocampus NMDA receptors differentially mediate trace and contextual fear conditioning.</a> Hippocampus. 15, 665-674 (2005).</li><li>McEchron, M. D., Bouwmeester, H., Tseng, W., Weiss, C., Disterhoft, J. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hippocampectomy+disrupts+auditory+trace+fear+conditioning+and+contextual+fear+conditioning+in+the+rat.">Hippocampectomy disrupts auditory trace fear conditioning and contextual fear conditioning in the rat.</a> Hippocampus. 8, 638-646 (1998).</li><li>Wiltgen, B. J., Sanders, M. J., Ferguson, C., Homanics, G. E., Fanselow, M. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Trace+fear+conditioning+is+enhanced+in+mice+lacking+the+delta+subunit+of+the+GABAA+receptor.">Trace fear conditioning is enhanced in mice lacking the delta subunit of the GABAA receptor.</a> Learn. Mem. 12, 327-333 (2005).</li><li>Davis, R. R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetic+basis+for+susceptibility+to+noise-induced+hearing+loss+in+mice.">Genetic basis for susceptibility to noise-induced hearing loss in mice.</a> Hear. Res. 155, 82-90 (2001).</li><li>Zheng, Q. Y., Johnson, K. R., Erway, L. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Assessment+of+hearing+in+80+inbred+strains+of+mice+by+ABR+threshold+analyses.">Assessment of hearing in 80 inbred strains of mice by ABR threshold analyses.</a> Hear. Res. 130, 94-107 (1999).</li><li>Moyer, J. R., Thompson, L. T., Disterhoft, J. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Trace+eyeblink+conditioning+increases+CA1+excitability+in+a+transient+and+learning-specific+manner.">Trace eyeblink conditioning increases CA1 excitability in a transient and learning-specific manner.</a> 16, 5536-5546 (1996).</li><li>Leuner, B., Falduto, J., Shors, T. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Associative+memory+formation+increases+the+observation+of+dendritic+spines+in+the+hippocampus.">Associative memory formation increases the observation of dendritic spines in the hippocampus.</a> J. Neurosci. 23, 659-665 (2003).</li><li>McEchron, M. D., Disterhoft, J. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hippocampal+encoding+of+non-spatial+trace+conditioning.">Hippocampal encoding of non-spatial trace conditioning.</a> Hippocampus. 9, 385-396 (1999).</li><li>McLaughlin, J., Skaggs, H., Churchwell, J., Powell, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Medial+prefrontal+cortex+and+pavlovian+conditioning:+trace+versus+delay+conditioning.">Medial prefrontal cortex and pavlovian conditioning: trace versus delay conditioning.</a> Behav. Neurosci. 116, 37-47 (2002).</li><li>Runyan, J. D., Moore, A. N., Dash, P. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+role+for+prefrontal+cortex+in+memory+storage+for+trace+fear+conditioning.">A role for prefrontal cortex in memory storage for trace fear conditioning.</a> J. Neurosci. 24, 1288-1295 (2004).</li><li>Gilmartin, M. R., McEchron, M. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Single+neurons+in+the+medial+prefrontal+cortex+of+the+rat+exhibit+tonic+and+phasic+coding+during+trace+fear+conditioning.">Single neurons in the medial prefrontal cortex of the rat exhibit tonic and phasic coding during trace fear conditioning.</a> Behav. Neurosci. 119, 1496-1510 (2005).</li><li>Crow, T., Xue-Bian, J. J., Siddiqi, V., Kang, Y., Neary, J. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Phosphorylation+of+mitogen-activated+protein+kinase+by+one-trial+and+multi-trial+classical+conditioning.">Phosphorylation of mitogen-activated protein kinase by one-trial and multi-trial classical conditioning.</a> J. Neurosci. 18, 3480-3487 (1998).</li><li>Martin, K. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=MAP+kinase+translocates+into+the+nucleus+of+the+presynaptic+cell+and+is+required+for+long-term+facilitation+in+Aplysia.">MAP kinase translocates into the nucleus of the presynaptic cell and is required for long-term facilitation in Aplysia.</a> Neuron. 18, 899-912 (1997).</li><li>Crestani, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Trace+fear+conditioning+involves+hippocampal+alpha5+GABA(A)+receptors.">Trace fear conditioning involves hippocampal alpha5 GABA(A) receptors.</a> Proc. Natl. Acad. Sci. U.S.A. 99, 8980-8985 (2002).</li><li>Crestani, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Decreased+GABAA-receptor+clustering+results+in+enhanced+anxiety+and+a+bias+for+threat+cues.">Decreased GABAA-receptor clustering results in enhanced anxiety and a bias for threat cues.</a> Nat. Neurosci. 2, 833-839 (1999).</li><li>Moore, M. D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Trace+and+contextual+fear+conditioning+is+enhanced+in+mice+lacking+the+alpha4+subunit+of+the+GABA(A)+receptor.">Trace and contextual fear conditioning is enhanced in mice lacking the alpha4 subunit of the GABA(A) receptor.</a> Neurobiol. Learn. Mem. 93, 383-387 (2010).</li><li>Cushman, J. D., Moore, M. D., Jacobs, N. S., Olsen, R. W., Fanselow, M. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Behavioral+pharmacogenetic+analysis+on+the+role+of+the+alpha4+GABA(A)+receptor+subunit+in+the+ethanol-mediated+impairment+of+hippocampus-dependent+contextual+learning.">Behavioral pharmacogenetic analysis on the role of the alpha4 GABA(A) receptor subunit in the ethanol-mediated impairment of hippocampus-dependent contextual learning.</a> Alcohol Clin. Exp. Res. 35, 1948-1959 (2011).</li><li>Raybuck, J. D., Lattal, K. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Double+dissociation+of+amygdala+and+hippocampal+contributions+to+trace+and+delay+fear+conditioning.">Double dissociation of amygdala and hippocampal contributions to trace and delay fear conditioning.</a> PLoS ONE. 6, (2011).</li><li>Kwapis, J. L., Jarome, T. J., Schiff, J. C., Helmstetter, F. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Memory+consolidation+in+both+trace+and+delay+fear+conditioning+is+disrupted+by+intra-amygdala+infusion+of+the+protein+synthesis+inhibitor+anisomycin.">Memory consolidation in both trace and delay fear conditioning is disrupted by intra-amygdala infusion of the protein synthesis inhibitor anisomycin.</a> Learn. Mem. 18, 728-732 (2011).</li><li>Gilmartin, M. R., Kwapis, J. L., Helmstetter, F. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Trace+and+contextual+fear+conditioning+are+impaired+following+unilateral+microinjection+of+muscimol+in+the+ventral+hippocampus+or+amygdala,+but+not+the+medial+prefrontal+cortex.">Trace and contextual fear conditioning are impaired following unilateral microinjection of muscimol in the ventral hippocampus or amygdala, but not the medial prefrontal cortex.</a> Neurobiol. Learn. Mem. 97, 452-464 (2012).</li><li>Baysinger, A. N., Kent, B. A., Brown, T. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Muscarinic+receptors+in+amygdala+control+trace+fear+conditioning.">Muscarinic receptors in amygdala control trace fear conditioning.</a> PLoS ONE. 7, (2012).</li><li>Wanisch, K., Tang, J., Mederer, A., Wotjak, C. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Trace+fear+conditioning+depends+on+NMDA+receptor+activation+and+protein+synthesis+within+the+dorsal+hippocampus+of+mice.">Trace fear conditioning depends on NMDA receptor activation and protein synthesis within the dorsal hippocampus of mice.</a> Behav. Brain. 157, 63-69 (2005).</li><li>Smith, D. R., Gallagher, M., Stanton, M. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetic+background+differences+and+nonassociative+effects+in+mouse+trace+fear+conditioning.">Genetic background differences and nonassociative effects in mouse trace fear conditioning.</a> Learn. Mem. 14, 597-605 (2007).</li><li>Rudy, J. W., O'Reilly, R. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Contextual+fear+conditioning,+conjunctive+representations,+pattern+completion,+and+the+hippocampus.">Contextual fear conditioning, conjunctive representations, pattern completion, and the hippocampus.</a> Behav. Neurosci. 113, 867-880 (1999).</li><li>Wiltgen, B. J., Sanders, M. J., Anagnostaras, S. G., Sage, J. R., Fanselow, M. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Context+fear+learning+in+the+absence+of+the+hippocampus.">Context fear learning in the absence of the hippocampus.</a> J. Neurosci. 26, 5484-5491 (2006).</li><li>Reijmers, L. G., Perkins, B. L., Matsuo, N., Mayford, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Localization+of+a+stable+neural+correlate+of+associative+memory.">Localization of a stable neural correlate of associative memory.</a> Science. 317, 1230-1233 (2007).</li><li>Huerta, P. T., Sun, L. D., Wilson, M. A., Tonegawa, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Formation+of+temporal+memory+requires+NMDA+receptors+within+CA1+pyramidal+neurons.">Formation of temporal memory requires NMDA receptors within CA1 pyramidal neurons.</a> Neuron. 25, 473-480 (2000).</li><li>Jacobs, N. S., Cushman, J. D., Fanselow, M. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+accurate+measurement+of+fear+memory+in+Pavlovian+conditioning:+Resolving+the+baseline+issue.">The accurate measurement of fear memory in Pavlovian conditioning: Resolving the baseline issue.</a> J. Neurosci. Methods. 190, 235-239 (2010).</li></ol>

Os autores não têm nada a revelar.

Houve vários estudos que têm elucidado o circuito neural que subjaz traço medo condicionado. Traçar medo condicionado é acreditado para envolver CA1 do hipocampo 12-14. Existe também evidência de que o córtex pré-frontal medial (CPFm) desempenha um grande papel no rastreio do pestanejar condicionado 15, e o CPFm foi encontrado para ser envolvido no rastreio de condicionamento de medo. Um estudo descobriu que os neurônios mPFC fornecer atividade sustentada durante o período de rastreio, p...

Com medo condicionado um estímulo neutro (NS) está emparelhado com um estímulo incondicionado aversivo (EUA). O NS é normalmente um tom e se torna um estímulo condicionado (CS) através de sorteios repetidos com os EUA. O CS pode então induzir uma resposta condicionada (CR), tal como a congelação, na ausência do aversivo EUA. Um protocolo de condicionamento do medo utilizada é atraso condicionado. Neste protocolo o início do NS e os EUA é contíguo ou com alguma sobreposição na apresentação do estímulo. Mesmo que demora medo condicionado é um dos tipos mais usados ​​de condicionamento associativo temporais, existem vários outros tipos de arranjos temporais condicionamento associativo: condicionamento simultâneo, condicionado para trás, e traço condicionado 1. No rastro do medo condicionado há um intervalo livre de estímulo entre o NS e os EUA de vários segundos, resultando em um período de &quot;trace&quot;. Vários estudos relataram déficits em rastreio de condicionamento de medo quando as lesões neurotóxicas são produzidas nas estruturas de entrada em que as hipocampo 2-5 ou quando os agentes farmacológicos são utilizados para bloquear a função do receptor no hipocampo. Lesão aos resultados hipocampo os défices traço e no condicionamento contextual, mas não prejudica atraso medo condicionado 8. Existem várias vantagens de se utilizar traço medo condicionado. O protocolo de condicionamento ao medo pode ser alcançado depois de um período de teste de três dias e permite a memória dependente do hipocampo que não é espacialmente dependente. Traçar medo condicionado pode ser usado como um exame complementar para o labirinto Morris água, romance teste de reconhecimento de objeto, ou outros testes de labirinto em investigar a memória dependente do hipocampo.

<table border="0" cellpadding="0" cellspacing="0" width="741">
	<colgroup>
		<col />
		<col />
		<col />
		<col />
	</colgroup>
	<tbody>
		<tr height="22">
			<td height="22" style="height:22px;width:216px;">FreezeFrame</td>
			<td style="width:143px;">Coulbourn</td>
			<td style="width:119px;"></td>
			<td style="width:263px;"></td>
		</tr>
		<tr height="41">
			<td height="41" style="height:41px;width:216px;">30% Isopropanol</td>
			<td style="width:143px;"></td>
			<td style="width:119px;"></td>
			<td style="width:263px;">Purchase 90% isopropanol and dilute it down to 30%</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">70% Ethanol</td>
			<td style="width:143px;"></td>
			<td style="width:119px;"></td>
			<td style="width:263px;"></td>
		</tr>
		<tr height="41">
			<td height="41" style="height:41px;width:216px;">Amp-meter</td>
			<td style="width:143px;">Med-Associates</td>
			<td style="width:119px;">ENV-420</td>
			<td style="width:263px;">Windows XP, Vista, and 7 Compatible (32-bit only)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Digital Sound Level Meter</td>
			<td style="width:143px;"></td>
			<td style="width:119px;">33-2055</td>
			<td style="width:263px;"></td>
		</tr>
		<tr height="41">
			<td height="41" style="height:41px;width:216px;">Vanilla Extract</td>
			<td style="width:143px;">McCormick Pure Vanilla Extract</td>
			<td style="width:119px;"></td>
			<td style="width:263px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Sticky Notes</td>
			<td style="width:143px;">Post-it</td>
			<td style="width:119px;"></td>
			<td style="width:263px;">3X3 inch</td>
		</tr>
	</tbody>
</table>

trace fear conditioning in mice

Neste experimento, apresentamos uma técnica para medir a aprendizagem ea memória. No protocolo condicionado traço medo aqui apresentada há cinco sorteios entre um estímulo neutro e um estímulo incondicionado. Há um período de rastreio de 20 segundos que separa cada ensaio condicionado. Na próxima congelamento dia é medido durante a apresentação do estímulo condicionado (CS) e período de vestígios. No terceiro dia, há um teste de 8 minutos para medir a memória contextual. Os resultados representativos são de camundongos que foram apresentados com o estímulo incondicionado aversivo (choque) em comparação com ratos que receberam as apresentações de tom, sem o estímulo incondicionado. Rastreamento medo condicionado foi usado com sucesso para detectar déficits e melhorias sutis de aprendizagem e memória em ratos que não são encontrados com outros métodos de medo condicionado. Este tipo de medo condicionado é considerada dependente da ligação entre o córtex pré-frontal e o hipocampo medial. Uma controvérsia atual é saber se este método é acreditado para ser independente de amígdala. Portanto, é necessário outro teste medo condicionado para examinar os efeitos de aprendizagem e memória dependente de amígdala, como através do atraso medo condicionado.

Para os resultados representativos são apresentados dados de ratos adultos da estirpe C57BL/6J de controlo que receberam os emparelhamentos de estímulo neutro com o estímulo não condicionado (choque condição) em comparação com os ratos que receberam o estímulo neutro, mas não receberam o estímulo não condicionado (nenhum estado de choque). É importante executar esta condição quando a primeira criação deste teste comportamental para determinar se o protocolo foi realizado corretamente. <p class="jove...

Watch this Scientific Journal Video about Trace Fear Conditioning in Mice at JoVE.com

Trace Fear Conditioning in Mice

No experimento seguinte, descrevemos um protocolo para traço medo condicionado em ratos. Este tipo de memória associativa inclui um período de traço que separa o estímulo neutro eo estímulo incondicionado.

Traçar medo condicionado em ratos

In this experiment we present a technique to measure learning and memory. In the trace fear conditioning protocol presented here there are five pairings ...

Research

JoVE Journal

Behavior

32.5K visualizações.  Baylor University.  No experimento seguinte, descrevemos um protocolo para traço medo condicionado em ratos. Este tipo de memória associativa inclui um período de traço que separa o estímulo neutro eo estímulo incondicionado. 

Vídeo: Trace Fear Conditioning in Mice

Contextual and Cued Fear Conditioning Test Using a Video Analyzing System in Mice

The contextual and cued fear conditioning test is one of the behavioral tests that assesses the ability of mice to learn and remember an association between environmental cues and aversive experiences. In this test, mice are placed into a conditioning chamber and are given parings of a conditioned stimulus (an auditory cue) and an aversive unconditioned stimulus (an electric footshock). After a delay time, the mice are exposed to the same conditioning chamber and a differently shaped chamber with presentation of the auditory cue. Freezing behavior during the test is measured as an index of fear memory. To analyze the behavior automatically, we have developed a video analyzing system using the ImageFZ application software program, which is available as a free download at http://www.mouse-phenotype.org/. Here, to show the details of our protocol, we demonstrate our procedure for the contextual and cued fear conditioning test in C57BL/6J mice using the ImageFZ system. In addition, we validated our protocol and the video analyzing system performance by comparing freezing time measured by the ImageFZ system or a photobeam-based computer measurement system with that scored by a human observer. As shown in our representative results, the data obtained by ImageFZ were similar to those analyzed by a human observer, indicating that the behavioral analysis using the ImageFZ system is highly reliable. The present movie article provides detailed information regarding the test procedures and will promote understanding of the experimental situation.

This article presents a protocol for a contextual and cued fear conditioning test using a video analyzing system to assess fear learning and memory in mice.

Contextual e Cued Medo Condicionado Teste Usando um sistema de análise de vídeo em Ratos

The contextual and cued fear conditioning test is one of the behavioral tests that assesses the ability of mice to learn and remember an association ...

Comportamento

Protocol for Studying Extinction of Conditioned Fear in Naturally Cycling Female Rats

Extinction of conditioned fear has been extensively studied in male rodents. Recently, there have been an increasing number of studies indicating that neural mechanisms for certain behavioral tasks and response behaviors are different in females and males. Using females in research studies can represent a challenge because of the variation of gonadal hormones during their estrous cycle. This protocol describes well-established procedures that are useful in investigating the role of estrogen in fear extinction memory consolidation in female rats. Phase of the estrous cycle and exogenous estrogen administration prior to extinction training can influence extinction recall 24 hr later. The vaginal swabbing technique for estrous phase identification described here aids the examination and manipulation of naturally cycling gonadal hormones. The use of this basic rodent model may further delineate the mechanisms by which estrogen can modulate fear extinction memory in females.

Gonadal hormones such as estrogen modulate memory formation in a number of experimental paradigms including fear extinction memory. This protocol describes a set of methods for investigating the influence of gonadal hormones specifically during extinction in naturally cycling females, including estrous cycle monitoring and exogenous estrogen administration.

Protocolo de Estudos de Extinção do medo condicionado em Naturally Ciclismo ratas

Extinction of conditioned fear has been extensively studied in male rodents. Recently, there have been an increasing number of studies indicating that ...

Quantifying Social Motivation in Mice Using Operant Conditioning

In this protocol, social motivation is measured in mice through a pair of operant conditioning paradigms. To conduct the experiments, two-chambered shuttle boxes were equipped with two operant levers (left and right) and a food receptacle in one chamber, which was then divided from the second chamber by an automated guillotine door covered by a wire grid. Different stimulus mice, rotated across testing days, served as a social stimulus behind the wire grid, and were only visible following the opening of the guillotine door. Test mice were trained to lever press in order to open the door and gain access to the stimulus partner for 15 sec. The number of lever presses required to obtain the social reward progressively increased on a fixed schedule of 3. Testing sessions ended after test mice stopped lever pressing for 5 consecutive minutes. The last reinforced ratio or breakpoint can be used as a quantitative measure of social motivation. For the second paradigm, test mice were trained to discriminate between left and right lever presses in order to obtain either a food reward or the social reward. Mice were rewarded for every 3 presses of each respective lever. The number of food and social rewards can be compared as a measurement of the value placed upon each reward. The ratio of each reward type can also be compared between mouse strains and the change in this ratio can be monitored within testing sessions to measure satiation with a given reward type. Both of these operant conditioning paradigms are highly useful for the quantification of social motivation in mouse models of autism and other disorders of social behavior.

This article describes a new protocol for the quantitative measurement of social motivation in mice using operant conditioning for a social reward. The protocol is useful for the measurement of social motivation in mouse models of autism and other disorders of social behavior.

Quantificar motivação social em ratos usando Condicionamento Operante

In this protocol, social motivation is measured in mice through a pair of operant conditioning paradigms. To conduct the experiments, two-chambered ...

Whisker-signaled Eyeblink Classical Conditioning in Head-fixed Mice

Eyeblink conditioning is a common paradigm for investigating the neural mechanisms underlying learning and memory. To better utilize the extensive repertoire of scientific techniques available to study learning and memory at the cellular level, it is ideal to have a stable cranial platform. Because mice do not readily tolerate restraint, they are usually trained while moving about freely in a chamber. Conditioned stimulus (CS) and unconditioned stimulus (US) information are delivered and eyeblink responses recorded via a tether connected to the mouse's head. In the head-fixed apparatus presented here, mice are allowed to run as they desire while their heads are secured to facilitate experimentation. Reliable conditioning of the eyeblink response is obtained with this training apparatus, which allows for the delivery of whisker stimulation as the CS, a periorbital electrical shock as the US, and analysis of electromyographic (EMG) activity from the eyelid to detect blink responses.

The preparation presented here for whisker-signaled eyeblink conditioning in head-fixed mice precisely stimulates specific whiskers while allowing mice to ambulate on a cylindrical treadmill. A whisker stimulation conditioned stimulus (CS) paired with a periorbital shock unconditioned stimulus (US) results in reliable associative learning on this apparatus.

sinalizado-Bigode Eyeblink condicionamento clássico em Ratos fixo-Cabeça

Eyeblink conditioning is a common paradigm for investigating the neural mechanisms underlying learning and memory. To better utilize the extensive ...

Reducing State Anxiety Using Working Memory Maintenance

The purpose of this protocol is to explain how to examine the relationship between working memory processes and anxiety by combining the Sternberg Working Memory (WM) and the threat of shock paradigms. In the Sternberg WM paradigm, subjects are required to maintain a series of letters in the WM for a brief interval and respond by identifying whether the position of a given letter in the series matches a numerical prompt. In the threat of shock paradigm, subjects are exposed to alternating blocks where they are either at risk of receiving unpredictable presentations of a mild electric shock or are safe from the shock. Anxiety is probed throughout the safe and threat blocks using the acoustic startle reflex, which is potentiated under threat (Anxiety-Potentiated Startle (APS)). By conducting the Sternberg WM paradigm during the threat of shock and probing the startle response during either the WM maintenance interval or the intertrial interval, it is possible to determine the effect of WM maintenance on APS.

This protocol demonstrates how to measure anxiety-potentiated startle during the Sternberg Working Memory paradigm.

Reduzindo a Ansiedade do Estado Usando a Manutenção da Memória de Trabalho

The purpose of this protocol is to explain how to examine the relationship between working memory processes and anxiety by combining the Sternberg Working ...

Classical Short-Delay Eyeblink Conditioning in One-Year-Old Children

Classical eyeblink conditioning (EBC) refers to the learned association between a conditioned stimulus (an auditory tone) and an unconditioned stimulus (a puff of air to the cornea). Eyeblink conditioning is often used experimentally to detect abnormalities in cerebellar-dependent learning and memory that underlies this type of associative learning. While experiments in adults and older children are relatively simple to administer using commercial equipment, eyeblink conditioning in infants is more challenging due to their poor compliance, which makes correct positioning of the equipment difficult. To achieve conditioning in one-year-old infants, a custom-made or an adapted commercial system can be used to deliver the air puff to the infant's cornea. The main challenge lies in successfully detecting and classifying the behavioral responses. We report that automated blink detection methods are unreliable in this population, and that conditioning experiments should be analyzed using frame-by-frame analysis of supplementary video camera recordings. This method can be applied to study developmental changes in eyeblink conditioning and to examine whether this paradigm can detect children with neurological disorders.

This protocol describes an eyeblink conditioning paradigm appropriate for experiments with one-year-old infants. Commercial or custom-made equipment can be used to deliver the stimuli, and data collection and analysis should be performed on the video recordings.

Condicionamento clássico curto-atraso Eyeblink em crianças de um ano de idade

Classical eyeblink conditioning (EBC) refers to the learned association between a conditioned stimulus (an auditory tone) and an unconditioned stimulus (a ...

Stress-Enhanced Fear Learning, a Robust Rodent Model of Post-Traumatic Stress Disorder

Fear behaviors are important for survival, but disproportionately high levels of fear can increase the vulnerability for developing psychiatric disorders such as post-traumatic stress disorder (PTSD). To understand the biological mechanisms of fear dysregulation in PTSD, it is important to start with a valid animal model of the disorder. This protocol describes the methodology required to conduct stress-enhanced fear learning (SEFL) experiments, a preclinical model of PTSD, in both rats and mice. SEFL was developed to recapitulate critical aspects of PTSD, including long-term sensitization of fear learning caused by an acute stressor. SEFL uses aspects of Pavlovian fear conditioning but produces a distinct and robust sensitized fear response far greater than normal conditional fear responses. The trauma procedure involves placing a rodent in a conditioning chamber and administering 15 unsignaled shocks randomly distributed over 90 minutes (for rat experiments; for mouse experiments, 10 unsignaled shocks randomly distributed over 60 minutes are used). On day 2, rodents are placed in a novel conditioning context where they receive a single shock; then, on day 3 they are placed back in the same context as on day 2 and tested for changes in freezing levels. Rodents that previously received the trauma display enhanced levels of freezing on the test day compared to those that received no shocks on the first day. Thus, with this model, a single highly stressful experience (the trauma) produces extreme fear of the stimuli associated with the traumatic event.

Here we describe the detailed methodology required to conduct stress-enhanced fear learning (SEFL) experiments, a preclinical model of post-traumatic stress disorder, in both rats and mice. The model utilizes aspects of Pavlovian fear conditioning and freezing as an index of enhanced fear in rodents.

Aprendizagem avançada stress medo, um modelo robusto de roedor do transtorno de estresse pós-traumático

Fear behaviors are important for survival, but disproportionately high levels of fear can increase the vulnerability for developing psychiatric disorders ...

Visual Classical Conditioning in Wood Ants

Several species of insects have become model systems for studying learning and memory formation. Although many studies focus on freely moving animals, studies implementing classical conditioning paradigms with harnessed insects have been important for investigating the exact cues that individuals learn and the neural mechanisms underlying learning and memory formation. Here we present a protocol for evoking visual associative learning in wood ants through classical conditioning. In this paradigm, ants are harnessed and presented with a visual cue (a blue cardboard), the conditional stimulus (CS), paired with an appetitive sugar reward, the unconditional stimulus (US). Ants perform a Maxilla-Labium Extension Reflex (MaLER), the unconditional response (UR), which can be used as a readout for learning. Training consists of 10 trials, separated by a 5-minute intertrial interval (ITI). Ants are also tested for memory retention 10 minutes or 1 hour after training. This protocol has the potential to allow researchers to analyze, in a precise and controlled manner, the details of visual memory formation and the neural basis of learning and memory formation in wood ants.

We present a protocol for the classical conditioning of harnessed ants that permits researchers to study visual learning and memory formation at a level of analysis not possible with freely moving individuals.

Visual condicionamento clássico em madeira formigas

Several species of insects have become model systems for studying learning and memory formation. Although many studies focus on freely moving animals, ...

Fear Incubation Using an Extended Fear-Conditioning Protocol for Rats

Emotional memory has been primarily studied with fear-conditioning paradigms. Fear conditioning is a form of learning through which individuals learn the relationships between aversive events and otherwise neutral stimuli. The most-widely utilized procedures for studying emotional memories entail fear conditioning in rats. In these tasks, the unconditioned stimulus (US) is a footshock presented once or several times across single or several sessions, and the conditioned response (CR) is freezing. In a version of these procedures, called cued fear conditioning, a tone (conditioned stimulus, CS) is paired with footshocks (US) during the training phase. During the first test, animals are exposed to the same context in which training took place, and freezing responses are tested in the absence of footshocks and tones (i.e., a context test). During the second test, freezing is measured when the context is changed (e.g., by manipulating the smell and walls of the experimental chamber) and the tone is presented in the absence of footshocks (i.e., a cue test). Most cued fear conditioning procedures entail few tone-shock pairings (e.g., 1-3 trials in a single session). There is a growing interest in less common versions involving an extensive number of pairings (i.e., overtraining) related to the long-lasting effect called fear incubation (i.e., fear responses increase over time without further exposure to aversive events or conditioned stimuli). Extended fear-conditioning tasks have been key to the understanding of fear incubation&#8217;s behavioral and neurobiological aspects, including its relationship with other psychological phenomena (e.g., post-traumatic stress disorder). Here, we describe an extended fear-conditioning protocol that produces overtraining and fear incubation in rats. This protocol entails a single training session with 25 tone-shock pairings (i.e., overtraining) and a comparison of conditioned freezing responses during context and cue tests 48 h (short-term) and 6 weeks (long-term) after training.

We describe an extended fear-conditioning protocol that produces overtraining and fear incubation in rats. This protocol entails a single training session with 25 tone-shock pairings (i.e., overtraining) and a comparison of conditioned freezing responses during context and cue tests 48 h (short-term) and 6 weeks (long-term) after training.

Incubação do medo usando um protocolo estendido de condicionamento do medo para ratos

Emotional memory has been primarily studied with fear-conditioning paradigms. Fear conditioning is a form of learning through which individuals learn the ...

Operant Conditioning Task to Measure Song Preference in Zebra Finches

An operant conditioning paradigm is used to test the song preference of female zebra finches. Finches are placed in a two-chambered cage with a connecting opening and indicate their preference for a song by landing on a perch within each chamber. By interrupting the infrared beam from a photoelectric sensor above each perch, the bird activates the playback of a song through a speaker located on each side of the cage. Freely available software is used to trigger the song playback from each perch. To determine the song preference of each animal, her chamber preference is first identified by triggering no song playback when she lands on each perch. This chamber preference is then compared to her song preference. A minimum activity threshold is set to ensure the preference is real. Using this method, we show that paired females prefer the song of their partner. This method was used to understand the contribution of dopamine to the formation and maintenance of song preference.

We describe a technique to evaluate song preference in zebra finches. Females are placed in a two-chambered cage and song preference is measured by the number of times she triggers the playback of one song by landing on a perch within one chamber, compared with triggering a different song in the second chamber. Perch landings are counted using infrared sensors.

Tarefa de condicionamento operante para medir a preferência da canção em tentilhões zebra

An operant conditioning paradigm is used to test the song preference of female zebra finches. Finches are placed in a two-chambered cage with a connecting ...